The efficiency and accuracy of the molecular mechanisms used by cells to repair DNA damaged by radiation play a crucial role in determining whether lesions will be properly mended or will lead to mutations, cancer or cell death. It is the major goal of this proposal to identify and understand the specific biological function of the genes and corresponding proteins involved in DNA repair. In order to address this problem, we will combine two eukaryotic systems, mammalian and the yeast Schizosaccharomyces pombe, and our approach will be three-fold. The first will include the isolation of human and/or mouse cDNAs capable of complementing the radiosensitivity of S. pombe rad3 and rad9 mutants, which are extremely sensitive to ionizing radiation and UV light, hypomutable and lack the ability to delay cycling in G2 following irradiation (i.e., molecular "checkpoint" control). Mammalian genes will be characterized with respect to structure, expression and other features. The second approach will involve the development of an in vitro DNA repair system, using S. pombe cell extracts. This system will be used to analyze the repair capability of S. pombe mutant cells, to identify mammalian and yeast proteins that have a common function and to purify repair proteins and complexes. The third goal will be to generate quantitative measures of radiation-induced DNA strand breaks to contribute to the formulation of models to account for the biological effects of radiation exposure. This combined molecular and biochemical strategy should help identify several genes and proteins involved in repair, aid in determining their specific function and define at least two genes that link DNA damage to cell cycle regulation.